Antibodies have long been used in medical diagnosis, e.g., determining blood types, and in biological experimentation. The usefulness of antibodies, however has been somewhat limited, as their complexity and diversity have made it very difficult to obtain homogeneous antibodies. Antibodies are complex protein or protein-based molecules which are produced by the immune systems of animals to protect the animal against foreign substances. Antibodies for medical use are generally obtained by injecting an animal with a foreign substance which will stimulate the animal's immune system and, most commonly, isolating an antibody fraction from the peripheral blood serum or from the ascitic fluid. The antibody fraction contains antibodies specific to the injected foreign substance as well as various other antibodies produced by the animal. By known techniques, it may be possible to substantially isolate an antibody specific to the particular foreign substance. However, even when an antibody for a particular foreign substance is isolated, such antibody is actually a mixture of several antibodies which recognize various antigenic determinants of the foreign substance or related substances. While some individual antibody molecules may be highly specific, recognizing only a certain foreign substance or portion thereof, other antibody molecules may be less selective, recognizing not only the subject foreign substance, but other substances as well. Because it is generally practically impossible to separate all related antibodies, even the most carefully purified antibody fractions may react with more than one substance.
In recent years, techniques of producing monoclonal antibodies (MAbs) have been developed which make it possible to obtain homogeneous, highly specific antibodies. Generally, such antibodies are produced by immunizing an animal with a protein fraction or other foreign substance, obtaining antibody-producing cells from the animal, and fusing the antibody-producing cells with strains of myeloma cells, e.g., tumor cells, to produce hybridomas which are isolated and cultured as monoclones. The monoclonal hybridomas may either be cultured in vitro or may be grown as tumors in a host animal. Because each antibody-producing cell produces a single unique antibody, the monoclonal cultures of hybridomas each produce homogeneous antibodies which may be obtained either from the culture medium of hybridoma cultures grown in vitro or from the cells, ascitic fluid, or serum of a tumor-bearing host animal.
Not all of the hybridoma clones which result from fusing neoplastic cells with antibody-producing cells are specific for the desired foreign substance or antigen (the substance with which the antibody reacts). This is because many of the hybridomas will make antibodies which the animal has produced to react with other foreign substances. Even antibodies against the subject antigen will differ from clone to clone because antibodies produced by different cells may react with different antigenic determinants of the same molecule. From each clone, therefore, it is necessary to obtain the resulting antibody or the antibody-containing medium, serum or ascitic fluid and test both its reactivity with the subject antigen and its specificity by determining what other substances, if any, it recognizes. While the necessity of characterizing the antibody of each clone adds to the complexity of producing monoclonal antibodies, the wide variety of homogeneous antibodies which may be obtained gives investigators a number of very precise tools to map the structure and development of somatic cells.
The availability of homogeneous, highly specific MAbs dramatically increases the value of antibodies as a diagnostic, experimental and therapeutic tool. Use of MAbs for tumor and virus detection has been described in U.S. Pat. Nos. 4,172,124 and 4,196,265.
MAbs are particularly suitable for studying the pathways and processes by which cells differentiate into different types of somatic cells to produce the various tissues of the body. Cell differentiation is a complex subject, and understanding of the processes involved is only beginning. Proteins which are specific to particular cell types and which may be detected by different MAbs, serve as precise markers for the study of cell development and differentiation. MAbs which are specific for given proteins not only may be used to ascertain the presence of known proteins in a cell, they may also be used to detect substances heretofore undiscovered. Theoretically it may be possible to eventually obtain MAbs for every macromolecule in the body to permit the complete mapping of the various proteins, etc.
An important topic in the field of cell differentiation is the study of cells which, in their mature form, are non-proliferating, being derived from actively proliferating stem cells. Many examples of such cells may be found in the peripheral blood. Red blood cells and leukocytes arise from stem cells in the bone marrow, and both are normally non-proliferating as mature cells in the blood stream. Misdevelopment of somatic cells may lead to cancers, including blood cell-related cancers such as myelomas and leukemias, and MAbs are useful for determining the proteins present in such cells to more fully trace their development and derivation.
In recent years, MAbs have been developed which react with the human transferrin (Tf) receptor. At least one of such MAbs blocks Tf binding to cells and thereby interferes with the ability of cells to proliferate. See, for example, U.S. Pat. No. 4,434,156, the disclosure of which is hereby incorporated by reference in its entirety. These MAbs are shown to be useful for inhibiting cell growth.
In Sauvage et al., Cancer Research, 47:747-753 (1987), two rat MAbs which bind to the murine Tf receptor are described which both block Tf receptor function and inhibit the growth of SL-2 leukemic cells in vitro when administered individually. In addition, the Sauvage article reports that one of these single anti-Tf MAbs, i.e., R17 208, in combination with an anti-Thy-1 MAb, which is directed to the Thy-1 glycoprotein and not to the murine Tf receptor, shows greater inhibitory effect against SL-2 leukemic cell growth in vitro. It then speculates that, based upon a test of only these two antibodies, it may be useful to use an anti-Tf antibody with one that activates a host immunological effector mechanism.
Despite the advances represented by these developments, there is always room for advancements which provide even more effective means for the regulation of cell growth.